Stepanenko Aleksei A, Sosnovtseva Anastasiia O, Valikhov Marat P, Vasiukova Anastasiia A, Abramova Olga V, Lipatova Anastasiia V, Yusubalieva Gaukhar M, Chekhonin Vladimir P
Department of Fundamental and Applied Neurobiology, V. P. Serbsky National Medical Research Center of Psychiatry and Narcology, Ministry of Health of the Russian Federation, 119034 Moscow, Russia.
Department of Medical Nanobiotechnology, Institute of Translational Medicine, N. I. Pirogov Russian National Research Medical University, Ministry of Health of the Russian Federation, 117997 Moscow, Russia.
Int J Mol Sci. 2025 Apr 14;26(8):3700. doi: 10.3390/ijms26083700.
The insertion of a transgene downstream of the L3-23K or L5-Fiber region was reported as a vital strategy for arming E3 non-deleted oncolytic adenoviruses. However, depending on the percentage of codons with G/C at the third base position (GC3%) and the type of splicing acceptor, an insert downstream of the L5-Fiber region may substantially affect virus fitness. Since the insertion of transgenes downstream of the L3-23K and L5-Fiber regions has never been compared in terms of their expression levels and impact on virus fitness, we assessed the total virus yield, cytolytic efficacy, and plaque size of Ad5-delta-24-RGD (Ad5Δ24RGD) armed with , , the suppressor of RNA silencing , soluble wild-type human/mouse and high-affinity human programmed cell death receptor-1 (PD-1/) ectodomains, and soluble human hyaluronidase PH20/. The insertion of transgenes downstream of the L3-23K region ensures their production at considerably higher levels. However, the insertion of transgenes downstream of either region differentially and unpredictably affects the oncolytic potency of Ad5Δ24RGD, which cannot be explained by GC3% or expression level alone. Surprisingly, the expression of the human and mouse PD-1 ectodomains with 83.1% and 70.1% GC3%, respectively, does not affect cytolytic efficacy but increases the plaque size in a cell line-dependent manner.
据报道，将转基因插入L3 - 23K或L5 - 纤维区域下游是武装E3未缺失的溶瘤腺病毒的关键策略。然而，根据第三碱基位置具有G/C的密码子百分比（GC3%）和剪接受体的类型，L5 - 纤维区域下游的插入可能会显著影响病毒适应性。由于从未在表达水平和对病毒适应性的影响方面对L3 - 23K和L5 - 纤维区域下游的转基因插入进行比较，我们评估了携带RNA沉默抑制因子、可溶性野生型人/小鼠和高亲和力人程序性细胞死亡受体-1（PD - 1/）胞外域以及可溶性人透明质酸酶PH20/的Ad5 - delta - 24 - RGD（Ad5Δ24RGD）的总病毒产量、细胞溶解效力和噬斑大小。在L3 - 23K区域下游插入转基因可确保其以相当高的水平产生。然而，在任一区域下游插入转基因会以不同且不可预测的方式影响Ad5Δ24RGD的溶瘤效力，这不能仅由GC3%或表达水平来解释。令人惊讶的是，人源和小鼠源PD - 1胞外域的表达，其GC3%分别为83.1%和70.1%，并不影响细胞溶解效力，但会以细胞系依赖的方式增加噬斑大小。
